Grzegorz Tylko

Bioactivity of coatings formed on Ti-13Nb-13Zr alloy using plasma electrolytic oxidation.

In this work, we investigated the bioactivity of anodic oxide coatings on Ti-13Nb-13Zr alloy by plasma electrolytic oxidation (PEO) in solutions containing Ca and P. The bioactive properties of the films were determined by immersion in simulated body fluid (SBF), and their biocompatibility was examined using adult human bone marrow derived mesenchymal stem cells (hBMSCs). The oxide layers were characterised based on their surface morphology (SEM, AFM, profilometry) as well as on their chemical and phase compositions (EDX, XRF, XRD, XPS). We report that anodic oxidation of Ti-13Nb-13Zr led to the development of relatively thick anodic oxide films that were enriched in Ca and P in the form of phosphate compounds. Furthermore, the treatment generated rough surfaces with a significant amount of open pores. The surfaces were essentially amorphous, with small amounts of crystalline phases (anatase and rutile) being observed, depending on the PEO process parameters. SBF soaking led to the precipitation of small crystals after one week of experiment. During culturing of hBMSCs on the bioactive Ti-13Nb-13Zr surfaces the differentiation of human mesenchymal stem cells toward osteoblasts was promoted, which indicated a potential of the modified materials to improve implant osseointegration.

Antifungal properties of silver nanoparticles against indoor mould growth

The presence of moulds in indoor environments causes serious diseases and acute or chronic toxicological syndromes. In order to inhibit or prevent the growth of microorganisms on building materials, the disruption of their vital processes or the reduction of reproduction is required. The development of novel techniques that impair the growth of microorganisms on building materials is usually based on silver nanoparticles (AgNPs). It makes them an alternative to other biocides. AgNPs have proven antibacterial activity and became promising in relation to fungi. The aim of the study was to assess growth and morphology of mycelia of typical indoor fungal species: Penicillium brevicompactum, Aspergillus fumigatus, Cladosporium cladosporoides, Chaetomium globosum and Stachybotrys chartarum as well as Mortierella alpina, cultured on agar media. The antifungal activity of AgNPs was also tested in relation to C. globosum and S. chartarum grown on the surface of gypsum drywall. It was found that the presence of AgNPs in concentrations of 30-200mg/l significantly decreased the growth of fungi. However, in the case of M. alpina, AgNPs stimulated its growth. Moreover, strong changes in moulds morphology and colour were observed after administration of AgNPs. Parameters of conidiophores/sporangiophores varied depending on mould region and changed significantly after treatment with AgNPs. The experiments have shown antifungal properties of AgNPs against common indoor mould species. Their application to building materials could effectively protect indoor environments from mould development. However, consideration must be given to the fact that the growth of some fungal strains might be stimulated by AgNPs.

Incorporation of sol-gel bioactive glass into PLGA improves mechanical properties and bioactivity of composite scaffolds and results in their osteoinductive properties.

In this study, 3D porous bioactive composite scaffolds were produced and evaluated for their physico-chemical and biological properties. Polymer poly-L-lactide-co-glycolide (PLGA) matrix scaffolds were modified with sol-gel-derived bioactive glasses (SBGs) of CaO-SiO2-P2O5 systems. We hypothesized that SBG incorporation into PLGA matrix would improve the chemical and biological activity of composite materials as well as their mechanical properties. We applied two bioactive glasses, designated as S2 or A2, differing in the content of SiO2 and CaO (i.e. 80 mol% SiO2, 16 mol% CaO for S2 and 40 mol% SiO2, 52 mol% CaO for A2). The composites were characterized for their porosity, bioactivity, microstructure and mechanical properties. The osteoinductive properties of these composites were evaluated in human bone marrow stromal cell (hBMSC) cultures grown in either standard growth medium or treated with recombinant human bone morphogenetic protein-2 (rhBMP-2) or dexamethasone (Dex). After incubation in simulated body fluid, calcium phosphate precipitates formed inside the pores of both A2-PLGA and S2-PLGA scaffolds. The compressive strength of the latter was increased slightly compared to PLGA. Both composites promoted superior hBMSC attachment to the material surface and stimulated the expression of several osteogenic markers in hBMSC compared to cells grown on unmodified PLGA. There were also marked differences in the response of hBMSC to composite scaffolds, depending on chemical compositions of the scaffolds and culture treatments. Compared to silica-rich S2-PLGA, hBMSC grown on calcium-rich A2-PLGA were overall less responsive to rhBMP-2 or Dex and the osteoinductive properties of these A2-PLGA scaffolds seemed partially dependent on their ability to induce BMP signaling in untreated hBMSC. Thus, beyond the ability of currently studied composites to enhance hBMSC osteogenesis, it may become possible to modulate the osteogenic response of hBMSC, depending on the chemistry of SBGs incorporated into polymer matrix.

Effects of Copper Supplementation on the Structure and Content of Elements in Kidneys of Mosaic Mutant Mice

Menkes disease is an effect of ATP7A gene mutation in humans, coding the Cu-ATP-ase which is essential in intestinal copper absorption and its subsequent transfer to circulation. This mutation results in a deficiency of copper in all tissues except the epithelia of intestine and kidney tubules. Subcutaneous injection of copper ions is the main therapy for Menkes patients. Mosaic (Atp7amo-ms) mice closely simulate the situation in Menkes disease. The aim of this study was to evaluate the changes in structure and element content in kidneys of mosaic mice after copper supplementation. Hematoxylin–eosin staining was used to analyze tissue morphology and atomic absorption spectrometry to estimate Cu and Zn content. X-ray microanalysis was performed to measure Na, Mg, P, Cl, and K content in the cells of the proximal and distal tubules. Copper administration lengthened the lifespan of the mutants but led to its high accumulation and results in severe kidney damage. Karyomegalia, necrosis of tubular and Bowman’s capsule epithelium, lesions, and atrophy of glomeruli were observed in the treated mutants. Copper treatment afterwards led to sclerosis of glomeruli and tubules enhanced proliferation of epithelial cells and formation of both polycystic and papillary carcinoma patterns in kidney. We suggest that copper excess may impair the activity of Na+/K+ ATP-ase in renal tubules of ms/− males. The content of Mg, P, and Cl in kidneys in mutants was also changed after copper administration.

Toxic effects of acrylamide on survival, development and haemocytes of Musca domestica

The influence of acrylamide, a potentially toxic substance present in some types of food, on survival, postembryonic development and haemocytes, insects blood cells, of the housefly was examined. Larvae were reared on media contaminated with acrylamide at concentrations of 82 microg/g, 164 microg/g or 246 microg/g. The length of larval and pupal stages as well as the survival of larvae and pupae was examined. To study the effects of acrylamide on haemocytes, the analysis of their index and morphology was performed in the third instar larva. The obtained data showed that the survival of larvae exposed to 82 microg/g and 164 microg/g concentrations of acrylamide decreased by 50% and 85%, respectively, whereas 246 microg/g concentration was lethal. In both groups of flies, larval and pupal stages were significantly lengthened by about 1.5 day in comparison with control. Moreover, acrylamide increased the number of prohaemocytes and intermediate cells while the number of plasmatocytes and granulocytes decreased. The size of plasmatocytes decreased in acrylamide-treated larvae when compared with these cells of control flies. The reduced survival of animals is probably due to affecting haemocytes involved in immune responses in insects. Moreover, the houseflys blood cells showed to be sensitive to toxin, which suggests their usefulness to test toxicity of substances present in food products.

Effects of zinc on programmed cell death of Musca domestica and Drosophila melanogaster blood cells.

Programmed cell death (PCD) and phagocytotic activity of immune cells play a pivotal role in insect development. We examined the influence of Zn(2+), an important element to fundamental biological processes, on phagocytosis and apoptosis of hemocytes in two fly species: Musca domestica and Drosophila melanogaster. Hemocytes were isolated from the third instar larvae of both species and treated for 3h with zinc chloride solutions, containing 0.35 mM or 1.7 mM of Zn(2+), and untreated as control. Phagocytotic activity of hemocytes was examined by flow cytometry after adding latex fluorescent beads to the medium, while apoptosis was evaluated by application of annexinV-FITC and pan-caspase-FITC inhibitor. Mitochondrial viability was determined by measuring resazurin absorbancy in the cell medium. The obtained results showed that Zn(2+) increases phagocytosis and affects PCD of both species hemocytes but each in a different way. Zinc decreases fraction of annexin-positive hemocytes in M. domestica but increases it in D. melanogaster. The pan-caspase analysis revealed low and high activity of caspases in hemocytes of M. domestica and D. melanogaster, respectively. Zn(2+) also decreased the viability of hemocyte mitochondria but only in D. melanogaster. It suggests that flies use different pathways of PCD, or that Zn plays a different role in this process in M. domestica than in D. melanogaster.

The effect of acrylamide and nitric oxide donors on human mesenchymal progenitor cells

We have examined the effects of nitric oxide donors and acrylamide on mesenchymal progenitor cell (hMPC) viability, programmed cell death (PCD) and differentiation. Acrylamide was examined at 0.5mM and 1.5mM concentrations, NOC-18 at 10μM and SNP at 100μM. Cell viability was assayed with MTS, PCD was determined by phosphatidylserine, caspase-9 and -3/7 and mitochondrial membrane potential assays, and osteogenic cell differentiation was evaluated by alkaline phosphatase activity (ALP) and mRNA levels for collagen type I, bone sialoprotein, ostepontin and osteocalcin. Serum-free hMPC cultures treated with 1.5mM acrylamide and SNP for 72h demonstrated reduced viability. PCD analyses revealed that SNP stimulated cells to necrosis in reactive species-dependent manner. Acrylamide (1.5mM) led to apoptosis independent of reactive species. Acrylamide and SNP reduced ALP activity and collagen type I mRNA levels but mRNA levels for bone sialoprotein and osteopontin increased in SNP treated cells and remained unchanged in acrylamide. Acrylamide had no effect on guanylate cyclase and cGMP osteogenic signaling pathway. The study suggests that acrylamide might impair bone development and remodeling upon acute or prolonged intoxication with this compound of mesenchymal cells.

Fibrinogen and carotid intima media thickness determine fibrin density in different atherosclerosis extents

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Elemental composition of Physarum compressum Alb. et Schw. sporocarps and their structures cultivated on rabbit dung and agar substrates

The elemental composition of spores, peridium walls, and lime nodes of Physarum compressum sporocarps, cultivated on rabbit dung as a natural growing environment for the slime mold and on artificial agar medium, was compared to evaluate differences that may be dependent on substrates. Whole fruiting bodies and samples of both experimental media were extracted with nitric acid or Parr digest bomb, respectively, and analyzed by means of total X‐ray reflection fluorescence (TXRF). Electron probe microanalysis (EPMA) of spores, peridium walls, and lime nodes structure was carried out with the scanning electron microscope equipped with energy‐dispersive spectrometer. Because of minute sizes and roughness of investigated structures, Monte Carlo simulations were utilized to establish analytical conditions of EPMA. Biological and geological standards were used in the quantification of element concentrations. According to TXRF, the fruiting bodies from agar medium revealed lower concentrations of K, Ca, Cr, Mn, and Fe in relation to fruiting bodies from the dung, reflecting elemental relationships in the experimental media. According to EPMA, the highest Ca concentration was found in the lime nodes followed by the peridium and the spores. Culturing of the slime molds on the rabbit dung indicated higher concentration of Ca in the lime nodes and peridium walls when compared with those obtained from the sporocarps grown on agar media. The opposite relation was found for the spores. The concentration of Na, Mg, P, S, and Cl was generally lower in all structures of the sporocarps harvested from the dung than from the agar medium. K was in higher concentration in analyzed structures from dung than from agar. Different element uptake (except for Ca and K) was revealed by the two methods: TXRF and EPMA. Microsc. Res. Tech. 73:1134–1142, 2010.

in vitro bioactivity investigations of Ti‐15Mo alloy after electrochemical surface modification

Titanium and its aluminum and vanadium-free alloys have especially great potential for medical applications. Electrochemical surface modification improves their surface bioactivity and stimulates osseointegration process. In this work, the effect of plasma electrolytic oxidation of the β-type alloy Ti-15Mo surface on its bioactivity is presented. Bioactivity of the modified alloy was investigated by immersion in simulated body fluid (SBF). Biocompatibility of the modified alloys were tested using human bone marrow stromal cells (hBMSC) and wild intestinal strains (DV/A, DV/B, DV/I/1) of Desulfovibrio desulfuricans bacteria. The particles of apatite were formed on the anodized samples. Human BMSC cells adhered well on all the examined surfaces and expressed ALP, collagen, and produced mineralized matrix as determined after 10 and 21 days of culture. When the samples were inoculated with D. desulfuricans bacteria, only single bacteria were visible on selected samples. There were no obvious changes in surface morphology among samples. Colonization and bacterial biofilm formation was observed on as-ground sample. In conclusion, the surface modification improved the Ti-15Mo alloy bioactivity and biocompatibility and protected surface against colonization of the bacteria.